Splitting hairs: Improved simultaneous detection of cocaine and alcohol use

Ezine

Published: Jul 23, 2013

Author: Steve Down

Channels: Gas Chromatography

Cocaine testing in hair

Hair testing for drugs of abuse is a growing area with distinct advantages over traditional urine analysis. The principal benefit to be gained is that a hair holds a historical record of drug use over time because the drugs or their metabolites remain unchanged once they enter the hair matrix. In spite of this, hair analysis is still regarded as a back up method, complementing urine testing.

In the case of cocaine, the parent drug and its principal metabolite benzoylecgonine (BE) are the principle targets to establish that a suspect has been taking this drug. If cocaethylene (CE) is also detected, that is an indication of the concurrent use of alcohol and cocaine, since it is produced in the liver by an enzyme-catalysed reaction between ethanol and cocaine. The presence of CE also rules out external contamination of hair as a means of defence as it is only produced within the body.

The main analytical techniques preferred for cocaine analysis in hair are immunoassays and mass spectrometry, specifically GC/MS and LC/MS. GC/MS is often the method of choice due to its sensitivity, selectivity and cost effectiveness but few methods have been reported for the simultaneous analysis of cocaine and CE.

Now, a GC/MS method has been introduced by Spanish scientists specifically targeted at patients undergoing dual alcohol and cocaine detoxification therapy. Olga López-Guarnido, Antonio Hernandez and colleagues from the University of Granada School of Medicine and the University of Santiago de Compostela described their procedure in Journal of Analytical Toxicology, in which they also propose a new criterion for minimizing false positive results.

Hair processing

The method involved the enzymatic digestion of hair, followed by solid-phase extraction and GC/MS with electron ionisation, a combination which has not been reported for the analysis of cocaine, CE and BE. Initially, the performance of the method was assessed using drug-free hairs that were spiked with known concentrations of the three analytes.

The samples were washed to remove any external contamination then cut into segments 1 mm long. These were heated with dithiothreitol to break down the disulphide bonds within the hair matrix, then digested with Pronase, which is a mixture of enzymes designed to break down completely the proteins in hair and release the small drug-related molecules.

Cocaine, BE and CE were removed from the resulting solution by SPE on a hydrophilic lipophilic balanced cartridge after adding the trideuterated analogues of the analytes to act as internal standards. The resulting extract was analysed by GC/MS using a 5% phenyl methylsilicone column to separate the drug components and selected ion monitoring to measure them.

The chromatogram displayed good separation of the three analytes with no interferences from other components present in the hair. It led to detection limits of 0.01, 0.03 and 0.04 ng/mg hair for cocaine, CE and BE, respectively, and the other analytical data like precision and accuracy were all acceptable.

Criteria for confirming cocaine use

The method was applied to hair samples taken from 40 cocaine users who had been arrested by police and admitted using the drug. They all tested positive for cocaine in urine tests.

Cocaine and BE were found in all of the hair samples. Levels of BE were on average 31.8% lower than those of cocaine and levels of the two compounds showed a positive correlation. The concentrations of cocaine ranged from 0.54 to 81.81 ng/mg, with amounts in the range 10-20 ng/mg being indicative of daily use. The metabolite CE was found in hair from 35 of the 40 subjects, with average amounts 4.8% those of cocaine. CE correlated positively with both cocaine and BE.

In order to completely rule out the possibility of external contamination by cocaine, it is generally recognised that three criteria should be fulfilled. The first two are a preliminary hair wash followed by detection of the appropriate metabolites in the hair. The third indicator relies on the establishment of cut-off levels for the analytes. So, if the amount of BE is more than 5% that of cocaine, cocaine use is regarded to be proven.

Now, this research team has proposed an additional condition, that the ratio of CE to cocaine should exceed 0.02. When this criterion was applied to the data from the 40 drug users, no false negatives were found. This is not surprising since all of the subjects were self-declared cocaine users. However, when the same criterion was applied to several sets of published data, it reduced the number of false negatives.

Even including this new factor, there is still no conclusive set of conditions which can be used to confirm cocaine use unambiguously by hair analysis, but it will improve the accuracy of the conclusions. It remains for organisations like the Society for Hair Testing and the Substance Abuse and Mental Health Services Administration to come up with recommendations which will improve testing and data interpretation so that no false results can be obtained.